PROJECT SUMMARY/ABSTRACT: Immunotherapy with monoclonal antibodies (mAbs) has been successful in settings such as autoimmunity and cancers, and therefore, treatment options with antibodies especially re-designed molecules based on broadly neutralizing antibodies (bNAbs) should be explored in the context of HIV-1. A caveat is that HIV-1 has the ability to rapidly escape from antibodies by generating mutations in its variable env gene. Therefore, there is an urgent need to gain insight into HIV-1 escape from bNAbs to aid in more effective combination antibody strategies to be used towards HIV-1 therapy, cure and prevention. As bNAbs are already being tested in clinical trials, it is imperative that optimal antibody combinations are evaluated not only for their neutralization capability but also their ?ease of escape? by diverse viruses. The overall objective of this proposal is to measure the ability of genetically diverse strains of HIV to escape from broadly neutralizing antibodies and design an antibody cocktail capable of restricting escape. To accomplish this goal, Dr. Lynch, who is an expert in HIV-1 escape from bNAbs has put together a collaborative team for an interdisciplinary approach using molecular virology (Dr. Lynch), computational methods (Drs. Barton and Fischer) and a humanized-mouse model (Dr. Klein) to study HIV-1 escape. Our central hypothesis is that HIV-1 escape from combination bNAbs will be limited when the mutations required to escape all bNAbs exert the maximum replicative fitness cost across diverse viruses and, therefore, replication cannot easily be restored through compensatory mutations. We will test this hypothesis by (i) defining a library of viable escape pathways for single bNAbs in diverse viruses with in vitro and in vivo approaches, (ii) determining the fitness cost or ?ease? of escape for virus-bNAb pairings bioinformatically, and (iii) identifying the optimal combination of antibodies that maximize fitness costs of resistance in diverse HIV-1 subtypes and testing this combination in vitro and in vivo. Our outcome will be the identification of optimal combination antibody cocktails to limit the ability of diverse HIV-1 viruses to escape from antibody pressure. These findings will inform all clinical trials using bNAbs or bNAb-based molecules, and ultimately, these studies will define a rational pipeline to characterize antibody escape pathways in the future.